It is estimated that approximately 218,000 Veterans were diagnosed with dementia in 2017, and 423,000 new cases of Alzheimer's disease and other dementias among military veterans in the decade ending in 2020. Alzheimer's disease (AD), Dementia with Lewy Bodies (DLB) and Parkinson's disease (PD) are all contributors to dementia which are associated with proteotoxicity; it is then critical to understand the mechanisms through which toxic proteins form and why their removal from neurons is inadequate. The link(s) between proteotoxicity, metabolism and the autophagy pathway, which removes toxic proteins, needs to be understood at a molecular level and in the context of neurodegenerative disease. This proposal focuses on an understudied pathway that is of interest in the sporadic degenerative diseases such as AD. In particular, perturbation of O-GlcNAcylation, as a nutrient sensing pathway, has been demonstrated in human Alzheimer's disease (AD) brains, and inhibition of O-GlcNAcase (OGA) has been shown effective in inhibiting tau phosphorylation in animal models. In light of these putative beneficial effects of increasing O-GlcNAc levels, clinical trials have been initiated to evaluate the efficacy of pharmacologically increasing this post-translational modification in treating AD. Whether the O- GlcNAcylation pathway is tau specific or also engages other neurotoxic proteins, such as ?-synuclein, has not been fully investigated in a mammalian system. We are among the first to demonstrate that O-GlcNAcylation is involved in ?-synuclein accumulation in vivo in mammalian systems and serves as the principal focus of this proposal. In support of this concept, we have shown that inhibition of OGA increased autophagosomal and ?- synuclein accumulation and attenuated autophagic flux in primary neurons. Our recent publication showing that that protein O-GlcNAcylation is increased in PD patients also underlines the growing need to understand the molecular regulation of this pathway. We have already developed the genetic animal models needed to test the overall hypothesis of this proposal that a sustained increase in O-GlcNAc causes ?-synuclein accumulation and neurological dysfunction, thereby contributing to the pathogenesis of AD and DLB. We have built a strong research team with expertise in O-GlcNAc biology, cutting-edge techniques in tag-mass spectrometry of the O-GlcNAc modification, autophagy, and mouse models of tissue specific OGA deficiencies and/or with ?-synuclein overexpression. This study will: 1) Test the hypotheses that increasing neuronal O- GlcNAc levels disrupts endogenous ?-synuclein homeostasis and impairs neurological function, 2) Test the hypothesis that O-GlcNAc modification of ?-synuclein as well as autophagy/endolysosomal proteins both contribute to ?-synuclein accumulation and thereby contributing to neurodegeneration. The successful completion of these studies will establish a role for O-GlcNAcylation in regulating ?-synuclein homeostasis and neurotoxicity, and provide new mechanistic understanding of O-GlcNAc regulation of autophagy, which will guide strategies for neuroprotection. For example, it is conceivable that introducing O-GlcNAcylation site peptide mimetic as competitive inhibitors may be beneficial to prevent detrimental O-GlcNAcylation while preserving beneficial O-GlcNAcylation. Success of this study may bring significant benefit to the Veterans who are affected by these diseases.